Systems and methods for transposing cargo nucleotide sequences

Abstract

The present disclosure provides systems and methods for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid. These systems and methods may comprise a double-stranded nucleic acid comprising the cargo nucleotide sequence, wherein the cargo nucleotide sequence is configured to interact with a recombinase or transposase complex, an effector complex comprising an effector and at least one engineered guide polynucleotide configured to hybridize to the target nucleic acid, and the recombinase or transposase complex wherein said recombinase or transposase complex is configured to recruit the cargo nucleotide to the target nucleic acid site.

What is claimed is: 
     
         1 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 2, type II Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide configured to hybridize to the target nucleic acid site;   b) a recombinase or transposase complex configured to bind the Cas effector complex; and   c) a double-stranded nucleic acid configured to interact with the recombinase or transposase complex and comprising the cargo nucleotide sequence.   
     
     
         2 . The system of  claim 1 , wherein the Cas effector complex binds non-covalently to the recombinase or transposase complex. 
     
     
         3 . The system of  claim 1 , wherein the Cas effector complex is covalently linked to the recombinase or transposase complex. 
     
     
         4 . The system of  claim 1 , wherein the Cas effector complex is fused to the recombinase or transposase complex. 
     
     
         5 . The system of any one of  claims 1-4 , wherein the cargo nucleotide sequence is flanked by a left-hand transposase recognition sequence and a right-hand transposase recognition sequence recognized by the recombinase or transposase complex. 
     
     
         6 . The system of  claim 5 , wherein the left-hand recombinase sequence comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 17-18. 
     
     
         7 . The system of any one of  claim 5 , wherein the right-hand recombinase sequence comprises a sequence having at least 80% identity to SEQ ID NO: 19. 
     
     
         8 . The system of  claim 1-7 , wherein the target nucleic acid comprises a PAM sequence compatible with the Cas effector complex. 
     
     
         9 . The system of  claim 8 , wherein the PAM sequence is located about 50 to about 70 base pairs from the target nucleic acid site. 
     
     
         10 . The system of  claim 9 , wherein the PAM sequence is located 3′ of the target nucleic acid site. 
     
     
         11 . The system of  claim 9 , wherein the PAM sequence is located 5′ of the target nucleic acid site. 
     
     
         12 . The system of any one of  claims 1-11 , wherein the class 2, type II Cas effector is not a Cas12k effector. 
     
     
         13 . The system of any one of  claims 1-11 , wherein the class 2, type II Cas effector comprises a polypeptide comprising a sequence having at least 80% identity to SEQ ID NO: 1. 
     
     
         14 . The system of any one of  claims 1-11 , wherein the class 2, type II Cas effector comprises a polypeptide comprising a sequence having at least 90% identity to SEQ ID NO: 1. 
     
     
         15 . The system of any one of  claims 1-11 , wherein the class 2, type II Cas effector comprises a polypeptide comprising a sequence of SEQ ID NO: 1. 
     
     
         16 . The system of any one of  claims 1-15 , wherein the recombinase or transposase complex comprises at least one polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 2-5. 
     
     
         17 . The system of any one of  claims 1-15 , wherein the recombinase or transposase complex comprises at least one polypeptide comprising a sequence having at least 90% identity to any one of SEQ ID NOs: 2-5. 
     
     
         18 . The system of any one of  claims 1-17 , wherein the recombinase or transposase complex comprises at least one polypeptide comprising a sequence of any one of SEQ ID NOs: 2-5. 
     
     
         19 . The system of any one of  claims 1-18 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to SEQ ID NO: 12. 
     
     
         20 . The system of any one of  claims 1-18 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 11. 
     
     
         21 . The system of any one of  claims 1-20 , wherein the small prokaryotic ribosomal protein subunit S15 comprises a sequence having at least 80% sequence identity to any one of any one of SEQ ID NOs: 494-659. 
     
     
         22 . The system of any one of  claims 1-21 , wherein the class 2, type II Cas effector and the recombinase or transposase complex are encoded by polynucleotide sequences comprising fewer than about 10 kilobases. 
     
     
         23 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 2, type V Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide configured to hybridize to the target nucleic acid site;   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising a TnsA, TnsB, TnsC, and TniQ component; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising the cargo nucleotide sequence.   
     
     
         24 . The system of  claim 23 , wherein the Cas effector complex binds non-covalently to the Tn7 type transposase complex. 
     
     
         25 . The system of  claim 23 , wherein the Cas effector complex is covalently linked to the Tn7 type transposase complex. 
     
     
         26 . The system of  claim 23 , wherein the Cas effector complex is fused to the Tn7 type transposase complex. 
     
     
         27 . The system of any one of  claims 23-26 , wherein the cargo nucleotide sequence is flanked by a left-hand transposase recognition sequence and a right-hand transposase recognition sequence recognized by the recombinase or transposase complex. 
     
     
         28 . The system of  claim 27 , wherein the left-hand recombinase sequence comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 20. 
     
     
         29 . The system of  claim 27 , wherein the right-hand recombinase sequence comprises a sequence having at least 80% identity to SEQ ID NO: 21. 
     
     
         30 . The system of any one of  claims 23-29 , wherein the target nucleic acid comprises a PAM sequence compatible with the Cas effector complex. 
     
     
         31 . The system of  claim 30 , wherein the PAM sequence is located about 50 to about 70 base pairs from the target nucleic acid site. 
     
     
         32 . The system of  claim 31 , wherein the PAM sequence is located 3′ of the target nucleic acid site. 
     
     
         33 . The system of  claim 31 , wherein the PAM sequence is located 5′ of the target nucleic acid site. 
     
     
         34 . The system of any one of  claims 23-33 , wherein the class 2, type V Cas effector is not a Cas12k effector. 
     
     
         35 . The system of any one of  claims 23-34 , wherein the TnsA component comprises a polypeptide comprising a sequence having at least 80% identity to SEQ ID NO: 7. 
     
     
         36 . The system of any one of  claims 23-25 , wherein the Tn7 type transposase complex comprises at least one polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 8-10. 
     
     
         37 . The system of any one of  claims 23-36 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to any one of SEQ ID NOs: 13-16. 
     
     
         38 . The system of any one of  claims 23-37 , wherein the small prokaryotic ribosomal protein subunit S15 comprises a sequence having at least 80% sequence identity to any one of any one of SEQ ID NOs: 494-659. 
     
     
         39 . The system of any one of  claims 23-38 , wherein the class 2, type II Cas effector and the recombinase or transposase complex are encoded by polynucleotide sequences comprising fewer than about 10 kilobases. 
     
     
         40 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 1, type I-F Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide configured to hybridize to the target nucleic acid site;   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising a TnsA, TnsB, TnsC, and TniQ component; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising a cargo nucleotide sequence.   
     
     
         41 . The system of  claim 40 , wherein the Cas effector complex binds non-covalently to the Tn7 type transposase complex. 
     
     
         42 . The system of  claim 40 , wherein the Cas effector complex is covalently linked to the Tn7 type transposase complex. 
     
     
         43 . The system of  claim 40 , wherein the Cas effector complex is fused to the Tn7 type transposase complex. 
     
     
         44 . The system of any one of  claims 40-43 , wherein the cargo nucleotide sequence is flanked by a left-hand transposase recognition sequence and a right-hand transposase recognition sequence recognized by the recombinase or transposase complex. 
     
     
         45 . The system of  claim 44 , wherein the left-hand recombinase sequence comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 136 and 138. 
     
     
         46 . The system of any one of  claim 44 , wherein the right-hand recombinase sequence comprises a sequence having at least 80% identity to SEQ ID NO: 137 and 139. 
     
     
         47 . The system of  claim 40-46 , wherein the target nucleic acid comprises a PAM sequence compatible with the Cas effector complex. 
     
     
         48 . The system of  claim 47 , wherein the PAM sequence is located about 50 to about 70 base pairs from the target nucleic acid site. 
     
     
         49 . The system of  claim 48 , wherein the PAM sequence is located 3′ of the target nucleic acid site. 
     
     
         50 . The system of  claim 48 , wherein the PAM sequence is located 5′ of the target nucleic acid site. 
     
     
         51 . The system of any one of  claims 40-50 , wherein the class 1, type I-F Cas effector comprises a polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 41-43 and 48-50. 
     
     
         52 . The system of any one of  claims 40-50 , wherein the class 1, type I-F Cas effector comprises a polypeptide comprising a sequence having at least 90% identity to any one of SEQ ID NOs: 41-43 and 48-50. 
     
     
         53 . The system of any one of  claims 40-50 , wherein the class 1, type I-F Cas effector comprises a polypeptide comprising a sequence of any one of SEQ ID NOs: 41-43 and 48-50. 
     
     
         54 . The system of any one of  claims 40-53 , wherein the Tn7 type transposase complex comprises at least one polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 44-47 and 51-54. 
     
     
         55 . The system of any one of  claims 40-53 , wherein the Tn7 type transposase complex comprises at least one polypeptide comprising a sequence having at least 90% identity to any one of SEQ ID NOs: 44-47 and 51-54. 
     
     
         56 . The system of any one of  claims 40-53 , wherein the Tn7 type transposase complex comprises at least one polypeptide comprising a sequence of any one of SEQ ID NOs: 44-47 and 51-54. 
     
     
         57 . The system of any one of  claims 40-56 , wherein the small prokaryotic ribosomal protein subunit S15 comprises a sequence having at least 80% sequence identity to any one of any one of SEQ ID NOs: 494-659. 
     
     
         58 . The system of any one of  claims 40-57 , wherein the class 2, type II Cas effector and the recombinase or transposase complex are encoded by polynucleotide sequences comprising fewer than about 10 kilobases. 
     
     
         59 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 22, 26, 30, 34, 55-89, 104, 147, 264-304, and 660-689; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 90-93, 111-114, 117, 151, 156-181, 201-204, 209-234, 255-258, 262, 263, 348, 350-353, 417-460, 491-492, and 715-739; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 23-25, 27-29, 31-33, 35-37, 101-103, 105-107, 148-150, 305-343, and 345-347; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 123, 125, 127, 129, 131, 133, 153, 354-358, 461, 463, 465, and 467; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 124, 126, 128, 130, 132, 154, 155, 359-363, 462, 464, 466, and 468. 
   
     
     
         60 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 22; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 90, 112, and 202; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 23-25; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 125; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 126 and 155. 
   
     
     
         61 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 26; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 91, 113, 156, 203, and 209; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 27-29; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 127; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 128. 
   
     
     
         62 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 60; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 117, 119, 161, and 214; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 101-103; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 131; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 132. 
   
     
     
         63 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 147; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 151, 181, and 234; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 148-150; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 153; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 154. 
   
     
     
         64 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site in a target nucleic acid and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 34; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 93, 114, 157, 204, and 210; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 148-150; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 129; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 130. 
   
     
     
         65 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 30; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 92, 111, and 201; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 31-33; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 123; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 124. 
   
     
     
         66 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex configured to hybridize to the target nucleic acid site and comprising:
 i) a class 2, type V Cas effector comprising a polypeptide having a sequence having at least 80% sequence identity to SEQ ID NO: 38; and 
 ii) an engineered guide polynucleotide comprising a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 98, 115-116, 182, 205-206, 235, and 493; 
   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB, TnsC, and TniQ components, the TnsB, TnsC, or TniQ component comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 39 and 40; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising in 5′ to 3′ order:
 i) a left-hand recombinase sequence comprising a sequence having at least 80% sequence identity to SEQ ID NO: 134; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase sequence comprising a sequence having at least 80% identity to SEQ ID NO: 135. 
   
     
     
         67 . The system of any one of  claims 59-66 , wherein the class 2, type V Cas effector is a Cas12k effector. 
     
     
         68 . The system of any one of  claims 59-67 , wherein the target nucleic acid comprises a PAM sequence compatible with the Cas effector complex. 
     
     
         69 . The system of  claim 68 , wherein the PAM sequence is located 5′ of the target nucleic acid site. 
     
     
         70 . The system of any one of  claims 68-69 , wherein the PAM sequence comprises 5′-nGTn-3′ or 5′-nGTt-3′. 
     
     
         71 . The system of any one of  claims 59-70 , wherein the Cas effector complex further comprises a small prokaryotic ribosomal protein subunit S15. 
     
     
         72 . The system of  claim 71 , wherein the small prokaryotic ribosomal protein subunit S15 comprises a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 494-659. 
     
     
         73 . The system of  claim 71 , wherein the class 2, type V Cas effector and the Tn7 type transposase complex are encoded by polynucleotide sequences comprising fewer than about 10 kilobases. 
     
     
         74 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 2, type V Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide configured to hybridize to the target nucleic acid site;   b) a Tn7 type transposase complex configured to bind the Cas effector complex and comprising TnsB and TnsC components but not a TnsA and/or TniQ component; and   c) a double-stranded nucleic acid configured to interact with the Tn7 type transposase complex and comprising the cargo nucleotide sequence.   
     
     
         75 . The system of  claim 74 , wherein the Cas effector complex binds non-covalently to the Tn7 type transposase complex. 
     
     
         76 . The system of  claim 74 , wherein the Cas effector complex is covalently linked to the Tn7 type transposase complex. 
     
     
         77 . The system of  claim 74 , wherein the Cas effector complex is fused to the Tn7 type transposase complex. 
     
     
         78 . The system of any one of  claims 74-77 , wherein the cargo nucleotide sequence is flanked by a left-hand transposase recognition sequence and a right-hand transposase recognition sequence recognized by the recombinase or transposase complex. 
     
     
         79 . The system of  claim 78 , wherein the left-hand recombinase sequence comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 134. 
     
     
         80 . The system of  claim 78 , wherein the right-hand recombinase sequence comprises a sequence having at least 80% identity to SEQ ID NO: 135. 
     
     
         81 . The system of any one of  claims 74-80 , wherein the target nucleic acid comprises a PAM sequence compatible with the Cas effector complex. 
     
     
         82 . The system of  claim 81 , wherein the PAM sequence is located about 50 to about 70 base pairs from the target nucleic acid site. 
     
     
         83 . The system of  claim 82 , wherein the PAM sequence is located 3′ of the target nucleic acid site. 
     
     
         84 . The system of  claim 82 , wherein the PAM sequence is located 5′ of the target nucleic acid site. 
     
     
         85 . The system of any one of  claims 74-84 , wherein the class 2, type V Cas effector is a Cas12k effector. 
     
     
         86 . The system of any one of  claims 74-85 , wherein the class 2, type V Cas effector comprises a polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 38 and 108. 
     
     
         87 . The system of any one of  claims 74-85 , wherein the class 2, type V Cas effector comprises a polypeptide comprising a sequence having at least 90% identity to any one of SEQ ID NOs: 38 and 108. 
     
     
         88 . The system of any one of  claims 74-85 , wherein the class 2, type V Cas effector comprises a polypeptide comprising a sequence of any one of SEQ ID NOs: 38 and 108. 
     
     
         89 . The system of any one of  claims 74-88 , wherein the TnsB subunit comprises a polypeptide comprising a sequence having at least 80% identity to SEQ ID NOs: 40 or 109. 
     
     
         90 . The system of any one of  claims 74-89 , wherein the TnsC subunit comprises a polypeptide comprising a sequence having at least 80% identity to SEQ ID NOs: 39 or 110. 
     
     
         91 . The system of any one of  claims 74-90 , wherein the Tn7 type transposase complex comprises at least one polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 39-40, 109-110, and 344. 
     
     
         92 . The system of any one of  claims 74-91 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 115, 116, 205, 206, 261, 235, 260, and 236. 
     
     
         93 . The system of any one of  claims 74-91 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to any one of SEQ ID NOs: 118, 182, 183, 235, and 236. 
     
     
         94 . The system of any one of  claims 74-93 , wherein the small prokaryotic ribosomal protein subunit S15 comprises a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 494-659. 
     
     
         95 . The system of any one of  claims 74-94 , wherein the class 2, type II Cas effector and the recombinase or transposase complex are encoded by polynucleotide sequences comprising fewer than about 10 kilobases. 
     
     
         96 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 2, type II Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide, the engineered guide polynucleotide capable of hybridizing to the target nucleic acid;   b) a recombinase or transposase complex operably linked to the Cas effector complex; and   c) a double-stranded nucleic acid comprising in 5′ to 3′ order:
 i) a left-hand recombinase recognition sequence; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase recognition sequence, the left-hand recombinase recognition sequence and the right-hand recombinase recognition sequence capable of being recognized by the recombinase or transposase complex. 
   
     
     
         97 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 2, type V Cas effector, a small prokaryotic ribosomal protein subunit S15, and an engineered guide polynucleotide, the engineered guide polynucleotide capable of hybridizing to the target nucleic acid;   b) a Tn7 type transposase complex operably linked to the Cas effector complex and comprising a TnsA, TnsB, TnsC, and TniQ component; and   c) a double-stranded nucleic acid comprising in 5′ to 3′ order:
 i) a left-hand recombinase recognition sequence; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase recognition sequence, the left-hand recombinase recognition sequence and the right-hand recombinase recognition sequence capable of being recognized by the Tn7 type transposase complex. 
   
     
     
         98 . A system for transposing a cargo nucleotide sequence into a target nucleic acid site in a target nucleic acid comprising:
 a) a Cas effector complex comprising a class 1, type I-F Cas effector, a small prokaryotic ribosomal protein subunit $15, and an engineered guide polynucleotide, the engineered guide polynucleotide capable of hybridizing to the target nucleic acid;   b) a Tn7 type transposase complex operably linked to the Cas effector complex and comprising a TnsA, TnsB, TnsC, and TniQ component; and   c) a double-stranded nucleic acid comprising in 5′ to 3′ order:
 i) a left-hand recombinase recognition sequence; 
 ii) the cargo nucleotide sequence; and 
 iii) a right-hand recombinase recognition sequence, the left-hand recombinase recognition sequence and the right-hand recombinase recognition sequence capable of being recognized by the Tn7 type transposase complex. 
   
     
     
         99 . An engineered nuclease system comprising:
 an endonuclease comprising a RuvC domain and an HNH domain, wherein the endonuclease is derived from an uncultivated microorganism, wherein the endonuclease is a Class 2, type II endonuclease comprising a sequence having at least 80% identity to SEQ ID NO: 1; and   an engineered guide polynucleotide, wherein the engineered guide RNA is configured to form a complex with the endonuclease and the engineered guide RNA comprises a spacer sequence configured to hybridize into a target nucleic acid sequence.   
     
     
         100 . The engineered nuclease system of  claim 99 , wherein the engineered guide polynucleotide comprises at least 60-80 consecutive nucleotides having at least 80% identity to SEQ ID NO: 12. 
     
     
         101 . The engineered nuclease system of  claim 99 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% identity to SEQ ID NO: 11. 
     
     
         102 . An engineered nuclease system comprising:
 an endonuclease comprising a RuvC domain, wherein the endonuclease is derived from an uncultivated microorganism, and wherein the endonuclease is a Class 2, type V endonuclease having at least 80% identity to SEQ ID NO: 6; and   an engineered guide RNA, wherein the engineered guide RNA is configured to form a complex with the endonuclease and the engineered guide RNA comprises a spacer sequence configured to hybridize into a target nucleic acid sequence.   
     
     
         103 . The engineered nuclease system of  claim 102 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to any one of SEQ ID NOs: 13-16. 
     
     
         104 . An engineered nuclease system comprising:
 an endonuclease comprising a RuvC domain, wherein the endonuclease is derived from an uncultivated microorganism, and wherein the endonuclease is a Class 2, type V-K endonuclease having at least 80% identity to any one of SEQ ID NOs: 22, 26, 30, 34, 55-89, 104, 147, 264-304, and 660-689; and   an engineered guide RNA, wherein the engineered guide RNA is configured to form a complex with the endonuclease and the engineered guide RNA comprises a spacer sequence configured to hybridize into a target nucleic acid sequence.   
     
     
         105 . The engineered nuclease system of  claim 104 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to any one of SEQ ID NOs: 90-93, 117, 151, 156-181, 209-234, 417-460, and 715-739. 
     
     
         106 . The engineered nuclease system of  claim 104 or 105 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% sequence identity to any one of SEQ ID NOs: 111-114, 201-206, 209, 210, 255-258, 262, 263, 348, 350-353, and 473-492. 
     
     
         107 . An engineered nuclease system comprising:
 an endonuclease comprising a RuvC domain, wherein the endonuclease is derived from an uncultivated microorganism, and wherein the endonuclease is a Class 2, type V-K endonuclease having at least 80% identity to SEQ ID NO: 38 or SEQ ID NO: 108; and   an engineered guide RNA, wherein the engineered guide RNA is configured to form a complex with the endonuclease and the engineered guide RNA comprises a spacer sequence configured to hybridize into a target nucleic acid sequence.   
     
     
         108 . The engineered nuclease system of  claim 107 , wherein the engineered guide polynucleotide comprises a sequence comprising at least about 46-80 consecutive nucleotides having at least 80% identity to any one of SEQ ID NOs: 118, 182, 183, 235, and 236. 
     
     
         109 . The engineered nuclease system of  claim 107 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 111-114, 115, 116, 201-206, 209, 210, 235, 236, 255-258, 260-263, 348, and 350-353. 
     
     
         110 . An engineered nuclease system comprising:
 a Class 1, type I-F Cas endonuclease comprising at least one Cas6, Cas7, or Cas8 polypeptide comprising a sequence having at least 80% identity to any one of SEQ ID NOs: 41-43 and 48-50; and   an engineered guide RNA, wherein the engineered guide RNA is configured to form a complex with the endonuclease and the engineered guide RNA comprises a spacer sequence configured to hybridize into a target nucleic acid sequence.   
     
     
         111 . The engineered nuclease system of  claim 110 , wherein the engineered guide polynucleotide comprises a sequence having at least 80% identity to any one of SEQ ID NOs: 121, 122, 207, and 208. 
     
     
         112 . A method for transposing a cargo nucleotide sequence into a target nucleic acid site comprising introducing the system of any one of  claims 1-111  to a cell. 
     
     
         113 . A cell comprising the system of any one of  claims 1-111 . 
     
     
         114 . The cell of  claim 113 , wherein the cell is a eukaryotic cell. 
     
     
         115 . The cell of  claim 113 , wherein the cell is a mammalian cell. 
     
     
         116 . The cell of  claim 113 , wherein the cell is an immortalized cell. 
     
     
         117 . The cell of  claim 113 , wherein the cell is an insect cell. 
     
     
         118 . The cell of  claim 113 , wherein the cell is a yeast cell. 
     
     
         119 . The cell of  claim 113 , wherein the cell is a plant cell. 
     
     
         120 . The cell of  claim 113 , wherein the cell is a fungal cell. 
     
     
         121 . The cell of  claim 113 , wherein the cell is a prokaryotic cell. 
     
     
         122 . The cell of  claim 113 , wherein the cell is an A549, HEK-293, HEK-293T, BHK, CHO, HeLa, MRC5, Sf9, Cos-1, Cos-7, Vero, BSC 1, BSC 40, BMT 10, WI38, HeLa, Saos, C2C12, L cell, HT1080, HepG2, Huh7, K562, primary cell, or a derivative thereof. 
     
     
         123 . The cell of  claim 113 , wherein the cell is an engineered cell. 
     
     
         124 . The cell of  claim 113 , wherein the cell is a stable cell.